Purpose :
To characterize in vitro corneas subjected to crosslinking with aqueous extract of Euterpe oleracea by enzymatic digestion using collagenase

Methods :
Rabbit eyes obtained from a slaughterhouse (Granja dos Ipês, Mairinque, Brazil) were deepithelialized and divided into three groups. The control group received no treatment. The eyes of the riboflavin/UVA group were treated according to standard procedure, receiving 0.1% riboflavin drops for 30 min and UV-A irradiation (365 nm) for 30 min (dose=5.4 J/cm2). The corneas subjected to açaí (Euterpe oleracea) were treated with 4% of the aqueous plant extract for 2 hours. The tissue was analyzed to determine the rate of enzymatic degradation by collagenase (Sigma-Aldrich) by overnight immersing in the DMEM/F12 medium (pH 7.4), containing dextran 10%, to achieve the hydration equilibrium and then submerged in collagenase 5.0 mg/ml. The mass values of the corneas were determined in different time intervals (0, 24, 48 h). The hydroxyproline content was measured in the supernatant at the initial time and after 24 h of digestion, using an assay kit (Sigma-Aldrich).

Results :
The rate of corneal digestion by collagenase was reduced in the crosslinked groups. The relative mass value in 24 and 48 h for the control group was 0.57 ± 0.09 and 0.32 ± 0.04, respectively. The corneas of the açaí group were not degraded by collagenase, and the relative mass values in 24 and 48 h were 1.02 ± 0.04 and 1.00 ± 0.03. In the riboflavin/UVA group, the mass had a statistically significant decrease (0.64 ± 0.13 in 24 h and 0.56 ± 0.16 in 48 h) which was lower than in the control group. The assay to determine the hydroxyproline content in 24 hours of digestion revealed that there was no digestion in the group treated with açaí. The values of the hydroxyproline concentration were 0.43 ± 0.04 mg/ml and 0.006 ± 0.0005 mg/ml for control and açaí groups, respectively. In both methodologies, the results clearly show that the açaí extract is adequate to avoid the biochemical degradation of corneas.

Conclusions :
The results of this work make clear that the corneas treated with Euterpe oleracea extract remained intact after enzymatic digestion by collagenase, which shows that corneal crosslinking using this plant extract was very effective

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.